The present invention relates to a modular apparatus for carrying out the essential phases of cell culture and analysis using robots if necessary.
The culture of procaryote and encaryote cells is practised in laboratories and industries in the main fields hereafter:
fundamental and applied research concerning the cell mechanisms (growth, differentiation, interaction) in varied disciplines (cell biology, cancerology, hormonology, study and testing of medicaments, etc . . . );
biotechnological research and industry with a view to cell manipulation and selection (hybridome clonage, for example):
toxicological analysis of foods, effluents, dyes or others, and
medical diagnosis passing through the preparation of human (cytogenetic, cancerology ) or bacterial cultures.
Cell culture is generally based on two kinds of methods, depending on the final result sought:
on the one hand, manual methods, widely used for small scale preparatory and analytic purposes, in which the main equipment hereafter is available:
thermostat controlled enclosures (with or without CO.sub.2),
laminarflow hoppers,
culture containers formed by Petri boxes, multihole boxes or flasks,
setting under culture or removal of cells or nutritive medium being carried out manually with a Pasteur pipette or, more recently, by means of semi-automatic devices with disposable plastic cones, whereas the morphological analysis generally takes place manually under an inverted microscope,
on the other hand, automated methods in bioreactors which allow the mass culture of floating cells for industrial production purposes and which are not directly concerned by the present invention.
It will be recalled that the manual methods comprise the main following phases:
1. Cell sampling (by dissociation,transplantation);
2. Placing under culture.,
3. Maintaining the cultures so as to obtain more especially growth and differentiation, which requires:
storage under defined temperature, hygrometry, 10% CO.sub.2 and pH conditions,
renewal of the nutritive medium, and
checking for non contamination and development of the process;
4. Interaction with the cell culture, for example by addition of an appropriate product or cells;
5. Taking cells for transplantation, or taking samples of the nutritive medium with a view to quantity determination, and
6. Morphological, biochemical, physical or similar analysis of the cell culture.
Said phases actually form a sequence of dissociated operations which, in all cases, leads to momentary variations in conditions all the consequences of which it is difficult to estimate. In fact, said culture containers are caused to transit between different apparatus:
incubator for storing the culture containers,
laminarflow hopper for carrying out the manipulations,
microscopic observation for checking or analysis purposes,
which leads at least to an increase in the risks of mutual contamination (man - culture cells) and also leads to a thermal shock which disturbs the cell metabolism.
In addition, the high number of culture containers to be maintained, the requirement of handling them as little as possible and the absence of adapted means prevent all the information contained in the cultures from being used.
To sum up, the main problems which arise with the manual methods at present used consist in:
high contamination risks with respect to cultivated cells (encaryotes especially), with respect to the personel if the cells are themselves pathogenic or if the manipulations involve dangerous agents (cancerogenic viruses, etc . . . );
need for a considerable staff force for carrying out work which is generally fastidious for the regular up-keep of the cell cultures, which results in considerable lost time due to the dispersion of the work posts and the difficulty of optimizing the operations;
high costs for specialized parts, consumable products (culture containers, nutritive media, etc . . . ) and for equipment, which should be multiplied so as to reduce the risks of contamination and so as to take into account the specialization of the application;
difficult biological working, related more particularly to the inevitable variability of human practice, to the non optimum environmental conditions (heat, mechanical and chemical shocks induced by handling), the impossibility of following and quantifying the biological processes which take place permanently in the cultures and a fbrtiori of acting retroactively thereon.
However, improvements have been made in some fields. Thus:
the incubator for storing the cultures and the laminarflow hopper have been juxtaposed by the firm HERAEUS;
numerous semi-automated devices exist for taking samples or diluting the cell medium;
constructors such as LEITZ propose microscopes contained in CO.sub.2 enclosures which are simply thermostat controlled but which do not provide sterility nor the constancy of a high degree of hygrometry, which can be explained in so far as the last point is concerned by the incompatibility with the optical and electromechanical components;
Prof. J.-C. BISCONTE has already proposed a device associating cell culture with automation, real-time and continuous image ana1ysis for following the cell interaction events (MIK-ROSKOPIE, 1980);
other solutions have been proposed, always in the direction of automation, such as those of WALKER and POZNANOVIC (International Biotechnology Laboratory, December 1983) who have constructed a multichamber device in which the conditions are programmable and which applies particulary to cells in suspension, but this device is not appropriate for:
the automated culture of a high number of samples,
the use of existing commercial containers,
morphological analysis,
automatic clonage and so is a device very different from that provided by the present invention.